# Development of CpG‐Depleted CFTR Plasmid‐Based Nanoparticles for Nonviral Gene Therapy in Lung Cystic Fibrosis Disease

## TL;DR

Researchers developed a new nonviral gene therapy approach for lung cystic fibrosis using CpG-depleted CFTR plasmid-based nanoparticles to achieve long-lasting protein expression.

## Contribution

A CpG-depleted and codon-optimized CFTR plasmid was developed to enable sustained and high-level CFTR protein expression.

## Key findings

- CpG-depleted CFTR plasmid nanoparticles achieved a 20-fold increase in CFTR protein expression in bronchial epithelial cells.
- The hEFIα promoter outperformed the CMV promoter, showing a 2.26-fold increase in CFTR expression at 7 days post-transfection.
- The plasmid-based approach demonstrated high efficacy in vitro, suggesting potential for in vivo application in cystic fibrosis treatment.

## Abstract

Nonviral gene therapy holds promise as a potential treatment for lung cystic fibrosis (CF). However, the transient expression of the CF transmembrane conductance regulator (CFTR) protein has limited its clinical application. To circumvent this challenge, a CpG‐depleted CFTR plasmid was developed. The CpG‐depleted CFTR plasmid could be compacted into DNA nanoparticles and modified with the addition of highly branched poly(β‐amino ester)s (HPAEs), leading to an improved and sustained CFTR protein expression. Using a CpG‐depleted and codon‐optimized CFTR sequence, around 20‐fold increase in CFTR protein production was achieved 48 h after treatment, compared with healthy human bronchial epithelial cells (16HBE14o‐). To evaluate the duration of CFTR protein expression induced by the plasmid based on human elongation factor 1α (hEFIα) and cytomegalovirus (CMV) promoters, a time course study was conducted in human CF bronchial epithelial (CFBE14o‐) cells. hEFIα promoter revealed a remarkable 2.26‐fold increase in CFTR protein expression at 7 days after transfection compared with 16HBE14o‐ cells. This level of CFTR protein expression outperformed the commonly used CMV promoter. The in vitro results demonstrated that CpG‐depleted CFTR plasmid could be used to achieve high efficacy in subsequent in vivo evaluations, which, if validated, may provide promising prospects for the development of a novel and effective treatment for lung cystic fibrosis.

Well designed CpG‐depleted CFTR nanoparticles for cystic fibrosis.